Transmembrane transport of Cl- will be studied to further characterize its role in two separate transport processes. In one, Cl- is a key participant (along with HCO3-, Na+ and H+) in a process that regulates intracellular pH (pHi), a function of fundamental importance in cellular physiology. In the other transport process to be studied, Cl is taken up along with Na+ and K+ into the squid giant axon. This process may be involved in cellular volume regulation, another fundamental cellular property. Experiments are proposed to determine the relative importance of ATP and the Na+ gradient as an energy source for the pHi regulatory system and for the Cl-:Na+:K+ uptake system. Additional studies on the pHi regulatory system are concerned with how the process is activated by a fall of pHi and whether or not Ca++ plays a role in this process. The current model for the pHi regulatory mechanism involves an external membrane-mediated exchange of external Na+ HCO3- for internal H+ Cl-. One of the goals of the present proposal is to test this model in some detail. The regulation of pHi is a relatively recent field of study and much remains to be learned about its ionic mechanism. The CL-Na+:K+ uptake system now identified in the squid giant axon has some similarities to systems described in other tissues, especially in the kidney tubules. The internally dialyzed giant axon provides a unique and powerful experimental advantage for the study of this transport system. Radioisotopic tracers and ion-selective microelectrodes will be used in these studies. The squid giant axon and the barnacle giant muscle fiber will be studied using the techniques of internal dialysis (for both preparations) and internal perfusion (barnacle giant muscle fibers). These cells were chosen specifically because their large size permits the use internal dialysis and perfusion techniques which allow experimental control of the intracellular environment and the precise measurement of unidirectional and net fluxes.